军曹鱼Hepcidin基因全长cDNA克隆及其组织表达分析

doi:10.16431/j.cnki.1671-7236.2015.07.002

›› 2015, Vol. 42 ›› Issue (7): 1630-1639.doi: 10.16431/j.cnki.1671-7236.2015.07.002

军曹鱼Hepcidin基因全长cDNA克隆及其组织表达分析

茅莉娜^1,2, 张鹤¹, 冯娟¹, 郭志勋¹, 许海东¹, 苏友禄¹

1. 中国水产科学研究院南海水产研究所, 农业部南海渔业资源开发利用重点实验室, 广州 510300;
2. 广东省疾病预防控制中心, 广州 511430

收稿日期:2014-12-17 出版日期:2015-07-20 发布日期:2015-07-23
通讯作者: 冯娟 E-mail:juanf@21cn.com
作者简介:茅莉娜(1984-),女,上海人,硕士,研究方向:动物病理生化及分子生物学,E-mail:49525894@qq.com;张鹤(1982-),女,河北张家口人,硕士,研究方向:鱼类分子免疫,E-mail:153024335@qq.com
基金资助:
国家自然基金青年基金(31202027)

Cloning of Full Length cDNA and Tissue Expression Analysis of Hepcidin Gene from Cobia (Rachycentron canadum)

MAO Li-na^1,2, ZHANG He¹, FENG Juan¹, GUO Zhi-xun¹, XU Hai-dong¹, SU You-lu¹

1. Key Laboratory of South China Sea Fishery Resources Exploitation and Utilization, Ministry of Agriculture, South China Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Guangzhou 510300, China;
2. Guangdong Provincial Center for Disease Control and Prevention, Guangzhou 511430, China

Received:2014-12-17 Online:2015-07-20 Published:2015-07-23

摘要/Abstract

摘要： 本试验采用同源克隆和末端快速扩增(RACE)的方法,得到714 bp的军曹鱼(Rachycentron canadum) Hepcidin基因全长cDNA序列。该序列包括213 bp的5'末端非编码区(UTR)、228 bp的3'UTR及273 bp的开放阅读框(ORF),编码90个氨基酸,预测其蛋白分子质量约为10.03 ku,等电点为7.54,预测的蛋白包括信号肽、前肽和成熟肽。通过构建Hepcidin氨基酸序列的系统进化树并进行氨基酸同源性比对,结果显示军曹鱼与已知鱼类及哺乳动物Hepcidin氨基酸的同源性在24.4%~85.6%之间。实时荧光定量PCR(quantitative Real-time PCR,qRT-PCR)检测结果显示Hepcidin基因在正常军曹鱼组织中均表达,但表达量在各个组织中有所不同,其中以肝脏表达量最高。经脂多糖(LPS)和甲醛灭活的鲨鱼弧菌(Vibrio carchariae)刺激后,肝脏、头肾、脾脏中Hepcidin基因表达均上调。

关键词: 军曹鱼; Hepcidin基因; 全长cDNA; 组织表达

Abstract: The techniques of homology clone and RACE were used to clone the Hepcidin gene from cobia (Rachycentron canadium).The full length cDNA of Hepcidin gene was 714 bp with a 213 bp 5'untranslated region (UTR),a 228 bp 3'UTR and a 273 bp open reading frame (ORF) encoding a polypeptide of 90 amino acid residues with a predicted molecular weight of 10.03 ku and theoretical isoelectric point of 7.54.The predicted molecular included signal peptide,prodomain peptide and mature peptide.Phylogenetic tree of Hepcidin amino acid sequences was constructed and homology compapison of amino acid sequences showed that homologies were varied from 24.4% to 85.6% with some known Hepcidin amino acids in other fishes and mammals.Quantitative Real-time PCR (qRT-PCR) analysis revealed that Hepcidin gene was expressed in all tissues with different expression levels,which expressed most in liver.The Hepcidin gene expressions in liver,head kidney and spleen were up-regulated after stimulation of LPS and formalin-inactivated Vibrio carchariae.

Key words: cobia (Rachycentron canadum); Hepcidin gene; full length cDNA; tissue expression

中图分类号:

茅莉娜, 张鹤, 冯娟, 郭志勋, 许海东, 苏友禄. 军曹鱼Hepcidin基因全长cDNA克隆及其组织表达分析[J]. , 2015, 42(7): 1630-1639.

MAO Li-na, ZHANG He, FENG Juan, GUO Zhi-xun, XU Hai-dong, SU You-lu. Cloning of Full Length cDNA and Tissue Expression Analysis of Hepcidin Gene from Cobia (Rachycentron canadum)[J]. , 2015, 42(7): 1630-1639.

参考文献

[1] Krause A,Neitz S,Magert H J,et al.LEAP-1,a novel highly disulfide-bonded human peptide,exhibits antimicrobial activity[J].Febs Lett,2000,480(2-3):147-150.
[2] Park C H,Valore E V,Waring A J,et al.Hepcidin,a urinary antimicrobial peptide synthesized in the liver[J].J Biol Chem,2001,276(11):7806-7810.
[3] 张书剑.几种鱼类抗菌肽的研究进展[J].饲料研究,2007,12:58-61.
[4] 周卫军,刘振兴,柯浩,等.鲢抗菌肽 Hepcidin 的基因克隆和表达及抑菌活性分析[J].南方水产科学,2014,10(3):58-64.
[5] Yang M,Wang K J,Chen J H,et al.Genomic organization and tissue-specific expression analysis of hepcidin-like genes from black porgy (Acanthopagrusschlegelii B)[J].Fish Shellfish Immunol,2007,23(5):1060-1071.
[6] Hirono I,Hwang J Y,Ono Y,et al.Two different types of hepcidins from the Japanese flounder Paralichthysolivaceus[J].FEBS J,2005,272(20):5257-5264.
[7] Douglas S E,Gallant J W,Liebscher R S,et al.Identification and expression analysis of hepcidin-like antimicrobial peptides in bony fish[J].Dev Comp Immunol,2003,27(6-7):589-601.
[8] Shike H,Shimizu C,Lauth X,et al.Organization and expression analysis of the zebrafish hepcidin gene,an antimicrobial peptide gene conserved among vertebrates[J]. Dev Comp Immunol,2004,28(7-8):747-754.
[9] 茅莉娜,冯娟,李玉谷,等.军曹鱼MHC-Ⅱα基因全长cDNA的克隆及其组织表达分析[J].中国畜牧兽医,2010,37(10):48-57.
[10] 王瑞旋,王江勇,徐力文,等.军曹鱼养殖水体及其肠道弧菌的耐药性研究[J].南方水产科学,2007,3(5):1-6.
[11] 李凌,吴灶和.鱼类体液免疫研究进展[J].海洋科学,2001,25(11):20-22.
[12] Bao B,Peatman E,Li P,et al.Catfish hepcidin gene is expressed in a wide range of tissues and exhibits tissue-specific upregulation after bacterial infection[J]. Dev Comp Immunol,2005,29(11):939-950.
[13] Bao B,Peatman E,Xu P,et al.The catfish liver-expressed antimicrobial peptide 2 (LEAP-2) gene is expressed in a wide range of tissues and developmentally regulated[J].Mol Immunol,2006,43(4):367-377.
[14] Courselaud B,Pigeon C,Inoue Y,et al.C/EBPalpha regulates hepatic transcription of hepcidin,an antimicrobial peptide and regulator of iron metabolism.Cross-talk between C/EBP pathway and iron metabolism[J].J Biol Chem,2002,277(43):41163-41170.
[15] Hilton K B,Lambert L A.Molecular evolution and characterization of hepcidin gene products in vertebrates[J]. Gene,2008,415(1-2):40-48.
[16] Chen S L,Xu M Y,Ji X S,et al.Cloning,characterization,and expression analysis of hepcidin gene from red sea bream (Chrysophrys major)[J].Antimicrob Agents Chemother,2005,49(4):1608-1612.
[17] 李伟,陈松林,张玉喜.牙鲆抗菌肽hepcidin基因的克隆及表达分析[J].高技术通讯,2007,17(1):78-82.
[18] 李伟,孙文秀.黄鳝Ⅰ型hepcidin抗菌肽基因的诱导表达分析[J].长江大学学报(自然科学版)农学卷,2009,2:52-55.
[19] Kim Y O,Hong S,Nam B H,et al.Molecular cloning and expression analysis of two hepcidin genes from olive flounder Paralichthys olivaceus[J].Biosci Biotechnol Biochem,2005,69(7):1411-1414.
[20] Shike H,Lauth X,Westerman M E,et al.Bass hepcidin is a novel antimicrobial peptide induced by bacterial challenge[J].Eur J Biochem,2002,269(8):2232-2237.
[21] 杨一新,李桂源.LPS所介导的信号转导通路研究进展[J].中南大学学报:医学版,2006,31(1):141-145.
[22] Lauth X,Babon J J,Stannard J A,et al.Bass hepcidin synthesis,solution structure,antimicrobial activities and synergism,and in vivo hepatic response to bacterial infections[J].J Biol Chem,2005,280(10):9272-9282.
[23] Wang K J,Cai J J,Cai L,et al.Cloning and expression of a hepcidin gene from a marine fish (Pseudosciaena crocea) and the antimicrobial activity of its synthetic peptide[J]. Peptides,2009,30(4):638-646.
[24] Chen S L,Li W,Meng L,et al.Molecular cloning and expression analysis of a hepcidin antimicrobial peptide gene from turbot (Scophthalmus maximus)[J].Fish Shellfish Immunol,2007,22(3):172-181.
[25] 郭明元,刘广锋,冯娟.1株军曹鱼病原弧菌的鉴定及其系统发育树分析[J].中国水产科学,2006,13(5):823-828.
[26] Collins J F,Wessling-Resnick M,Knutson M D.Hepcidin regulation of iron transport[J]. J Nutr,2008,138(11):2284-2288.
[27] Nemeth E,Tuttle M S,Powelson J,et al.Hepcidin regulates cellular iron efflux by binding to ferroportin and inducing its internalization[J].Science,2004,306(5704):2090-2093.
[28] 牛诗雯,宁波,刘罡,等.Hepcidin与疾病关系和其临床意义的研究进展[J].生命科学,2012,8:817-826.

军曹鱼Hepcidin基因全长cDNA克隆及其组织表达分析

Cloning of Full Length cDNA and Tissue Expression Analysis of Hepcidin Gene from Cobia (Rachycentron canadum)

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